T cell-based therapies have limited success against ovarian cancer for poorly understood reasons. Here we show that lipids in ovarian cancer ascites fluid disrupt nanoscale T cell receptor dynamics-driven T cell activation. T cells stimulated in ascites of patients with ovarian cancer have an altered lipid profile, including changes in phosphatidylcholine (PC) and phosphatidylethanolamine content, and disrupted membrane properties. Using untargeted lipidomics, we reveal which lipid species are consumed by T cells in ascites and show that 18:0-18:2 PC impairs T cell activation at physiological concentrations. Mechanistically, we uncover that lipids in ascites, including 18:0-18:2 PC, disrupt nanoscale T cell receptor clustering in immunological synapses. Importantly, pre-activated T cells overcome these lipid-induced barriers, highlighting a promising strategy to enhance adoptive T cell-based therapies for ovarian cancer. These findings provide mechanistic insights into lipid-mediated immune suppression in the tumour microenvironment and an actionable strategy to overcome these barriers.